Peroxisome biogenesis disorders (PBD) affect 1/50,000 births and range in severity from death to developmental delays and metabolic disruptions in brain, liver, and kidneys. Peroxisomes are small organelles found within all cells that contain an important subset of metabolic reactions and are essential for normal growth and development. PBD cells contain fewer peroxisomes and those present often have no metabolic activity. Disease phenotypes can be rescued by increasing peroxisomal functions, as symptoms result from decreased activity. Plant peroxisomes similarly are essential. Mutants defective in peroxisomal processes have delayed development, due to decreased metabolism, and do not make required plant hormones, resulting in altered plant architecture and reduced fertility. Several metabolic engineering projects for increasing plant health or improving human nutrition have been unsuccessful due to activation of peroxisomal metabolism that limits product accumulation. However, these projects have shown promise when done in backgrounds with decreased peroxisomal activity. Peroxisomal processes occur routinely during normal growth, with increased activity during developmental stages or in environmental conditions that require a higher level of peroxisomal products. This proposal will address how peroxisomal pathways are regulated, increasing our ability to influence activity in situations where metabolism is problematically low (peroxisomal biogenesis disorders) or high (engineering plants to generate novel products). We will first examine how peroxisomal pathways are regulated at both the gene and the protein level by comparing phenotypic responses between normal plants and mutant lines with reduced function or altered regulation. Second, we will examine whether all peroxisomes contain the same complement of enzymes or if individual peroxisomes have specialized activity. We will further examine how peroxisomal pathways regulate each other. Finally, we will investigate how peroxisomal metabolism influences peroxisome size. Peroxisomes are larger in mutants with disrupted processing, presumably as a feedback response to increase activity. We will examine how metabolism affects size and characterize the signal(s) initiating the feedback loop that links required pathway outputs with organelle growth. This proposal serves the dual purpose of providing information about peroxisomal functions that can be applied broadly in other organisms as well as exploring specific mechanisms required for plant growth, development, and responses to a changing environment.